This project is part of a long-term collaborative effort between this laboratory and Dr. Shyh-Ching Lo?s lab at AFIP to investigate the co-factors contributing to the pathogenesis of AIDS. It has been a fruitful scientific and intellectual collaboration. The laboratory continues to support the work on diagnosis and characterization of mycoplasma originating from AIDS patients, sexually transmitted diseases (STDs) patients, and others. We applied serological tests that were developed in this laboratory to patients in several clinical settings, including patients with HIV infection, nongonococcal urethritis (NGU), STDs, and intravenous drug use. We found a high prevalence of antibodies to M. penetrans in patients with Kaposi?s sarcoma and antibodies to M. genitalium in patients with NGU. Using the paired donor-recipient specimens, we also found that M. fermentans and M. genitalium were transmissible through blood transfusion. We were able to show that the association of the presence of antibodies to M. genitalium with the sexual transmission of HIV was highly significant, while agents for other STDs were not. In recent years, we were asked to support Dr. Lo?s lab by providing serological tests on specimens from patients who suffered from the Gulf War syndrome or Gulf War infection (GWI). Over 6,000 paired specimens, including controls, were examined. We were not able to find any difference between soldiers who served in the Gulf War and their controls in antibody seroconversion to M. fermentans. To clarify a report that more than 50 percent of veterans with GWI had M. fermentans (strain incognitus) in their blood as measured by a molecular diagnostic technique called nuclear gene tracking, we conducted a large-scale, case-control study to compare the prevalence of antibodies to M. fermentans lipid-associated membrane proteins (LAMPs) between the Gulf War veterans with unexplained illness and a randomly selected, matched group of veterans who did not enroll in the registry for health evaluation. In addition, we analyzed, using banked serum samples obtained on each individual before and after the deployment, the rates of seroconversion for this mycoplasma in these two groups of veterans. Our results showed 4.8 percent of the cases and 5.2 percent of the controls tested positive for M. fermentans-specific antibodies before operation deployment. Most important, there was no difference in rates of seroconversion between cases and controls (1.1 vs. 1.2 percent) to M. fermentans during ODS. Thus, there is no serological evidence that suggests infection by M. fermentans is associated with development of GWI. We also studied blood, urine, oral swabs, and rectal swabs for evidence of mycoplasmal infection by culture from a group of 149 Gulf War veterans who complained of various illnesses and were enrolled in the second phase of the health evaluation by the Army Comprehensive Clinical Examination Program (CCEP). None of the urine samples, oral swabs, or rectal swabs grew M. fermentans. No mycoplasma organism was isolated from any of the 149 blood samples. PCR study was conducted using RW oligonucleotide primer set (RW004 and RW005), based on the unique sequence of the M. fermentans insertion-sequence-like element. The amplified products were confirmed by southern blot using RW006 as the hybridization probe. Each sample was tested in triplicate at least three times. Three out of 65 (4 percent) blood samples were considered positive. Two of these three patients tested positive for M. fermentans antibodies in the serological study. In conclusion, our culture study of ODS veterans with GWI revealed isolation of only mycoplasma organisms commonly found in similar samples from healthy individuals. No unusual mycoplasma was identified. Contrary to reported studies from some other laboratories, our PCR and serological studies showed only a low percentage of the veterans having evidence of M. fermentans infection. One of the future interests for mycoplasma study is its contribution to the development of neoplasms after long-term, low-level chronic infection. We have shown that some species of mycoplasma were able to transform cells in vitro after long-term co-cultivation, and several indicative oncogenes were activated. We are continue our collaboration but limited our effort to the advising and technical support level.